title: Development of an autofluorescent whole-cell biocatalyst by displaying dual functional moieties on Escherichia coli cell surfaces and construction of a coculture with organophosphate-mineralizing activity .
abstract: Surface display of the active proteins on living cells has enormous potential in the degradation of numerous toxic compounds. Here, we report the codisplay of organophosphorus hydrolase (OPH) and enhanced green fluorescent protein (GFP) on the cell surface of Escherichia coli by use of the truncated ice nucleation protein (INPNC) and Lpp-OmpA fusion systems. The surface localization of both INPNC-OPH and Lpp-OmpA-GFP was demonstrated by Western blot analysis, immunofluorescence microscopy, and a protease accessibility experiment. Anchorage of GFP and OPH on the outer membrane neither inhibits cell growth nor affects cell viability, as shown by growth kinetics of cells and stability of resting cultures. The engineered E. coli can be applied in the form of a whole-cell biocatalyst and can be tracked by fluorescence during bioremediation. This strategy of codisplay should open a new dimension for the display of multiple functional moieties on the surface of a bacterial cell. Furthermore, a coculture comprised of the engineered E. coli and a natural p-nitrophenol (PNP) degrader, Ochrobactrum sp. strain LL-1, was assembled for complete mineralization of organophosphates (OPs) with a PNP substitution. The coculture degraded OPs as well as PNP rapidly. Therefore, the coculture with autofluorescent and mineralizing activities can potentially be applied for bioremediation of OP-contaminated sites.
authors.affiliation: State Key Laboratory of Integrated Management of Pest Insects & Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Yang
authors.forename: Yaran
authors.initials: Y
authors.lastname: Zhu
authors.forename: Jijian
authors.initials: J
authors.lastname: Yang
authors.forename: Zheng
authors.initials: Z
authors.lastname: Liu
authors.forename: Chuanling
authors.initials: C
authors.lastname: Qiao
authors.forename: Ashok
authors.initials: A
authors.lastname: Mulchandani
authors.forename: Wilfred
authors.initials: W
authors.lastname: Chen
mesh_terms: D043303:Aryldialkylphosphatase / Q000235:genetics / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D018920:Coculture Techniques; D004798:Enzymes / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D050296:Microbial Viability; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009596:Nitrophenols / Q000378:metabolism; D042041:Ochrobactrum / Q000235:genetics / Q000378:metabolism; D010755:Organophosphates / Q000378:metabolism*; D010447:Peptide Hydrolases / Q000378:metabolism; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA
chemical_list: D001425:Bacterial Outer Membrane Proteins; D004798:Enzymes; D029968:Escherichia coli Proteins; D009596:Nitrophenols; D010755:Organophosphates; D012336:RNA, Ribosomal, 16S; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D043303:Aryldialkylphosphatase; D010447:Peptide Hydrolases; C024836:4-nitrophenol